Structure-Activity Relationship in Host Defense Peptides

宿主防御肽的构效关系

基本信息

批准号：
7940023
负责人：
Gregory Alexander Caputo
金额：
$ 29.24万
依托单位：
ROWAN UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-07-01 至 2013-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7940023
关键词：
Affect Affinity Amino Acids Amphibia Antibiotic Resistance Antibiotics Bacteria Behavior Binding Biological Assay Biological Models Cell Membrane Permeability Cell membrane Cell surface Cells Cessation of life Charge Chemicals Circular Dichroism Circular Dichroism Spectroscopy Data Dependence Detergents Development Energy Transfer Environment Environmental Risk Factor Exhibits Exposure to Fluorescence Fluorescence Spectroscopy Gene Mutation Goals Gram-Positive Bacteria Hospitals Host Defense Hydrophobicity Immune system Insecta Investigation Ionic Strengths Lateral Link Lipid Bilayers Lipids Measures Membrane Membrane Lipids Membrane Proteins Methods Micelles Modeling Modification Optics Organism Parents Patients Penetration Peptide Synthesis Peptides Plasmids Prevalence Property Proteins Public Health Recombinant Proteins Research Research Project Grants Resistance Resistance development Resolution Role Screening procedure Series Site Source Specificity Spectrum Analysis Staging Structure Structure-Activity Relationship Surface System Techniques Testing Vesicle X-Ray Crystallography analog antimicrobial antimicrobial peptide aqueous base design emission spectroscopy environmental change flexibility fluorophore in vivo insight membrane model pancreatic secretory trypsin inhibitor I pathogen peptide structure protein aminoacid sequence protein folding protein protein interaction public health relevance research study response restraint small molecule

项目摘要

DESCRIPTION (provided by applicant): The overall goal of this research is to chemical and structural motifs linked to the mechanism of action of membrane-interacting antimicrobial peptides. The increased prevalence of antibiotic resistance in numerous bacterial strains has a tremendous annual impact, especially in hospital settings for patients with compromised immune systems. As more strains develop resistance to more antibiotics it underscores the need for development of new antimicrobials that have a low propensity for resistance development. One potential source of such antimicrobials are naturally occurring host-defense peptides (HDPs) which have been found in a variety of organisms throughout the evolutionary chain, indicative that bacteria have not been able to mount a sufficient resistance response to these molecules. The HDPs are typically short peptides (10-50aa) that bind to bacterial cell surfaces with high affinity and high selectivity over host cells. The mechanism of action of the peptides is still a matter of debate, but most HDPs exhibit some degree of membrane destabilization upon binding to the lipid bilayer which is likely linked to the mechanism of action. Recent studies have also shown a number of bacterial transcriptional responses induced by exposure to HDPs, many of which are not completely understood. Mechanism of action studies are further complicated by the sheer number of HDPs isolated from natural sources, many of which may use somewhat different mechanisms based on the host and what pathogens the host routinely encounters. The goal of the proposed research is to focus on the mechanism of action of several naturally derived alpha- helical HDPs selected for diverse host organisms and broad spectrum antimicrobial activity. A deeper understanding of the mechanism of alpha-helical HDPs relies on understanding the structure-activity relationship in these peptides. Furthermore, as relatively small peptides, the HDPs are experimentally tractable and valuable biophysical model for understanding protein-membrane interaction and as a model system for exploring the principles of membrane protein folding. Our strategy for studying the behavior of HDPs involves the synthesis of peptides with natural and altered peptide sequences designed to examine the role(s) of specific chemical moieties in the activity of the peptide. This will be achieved by rationally incorporating either naturally occurring or synthetic amino acids to replace similar residues at a series of sites within the peptide, monitoring the binding of modified peptides to lipid bilayers, and testing the antimicrobial activity of the newly designed peptides. Multiple types of optical spectroscopy will be employed to characterize the membrane binding and structural transitions in the peptides. Fluorescence emission spectroscopy is a sensitive probe of environmental changes (i.e. aqueous peptide binding to a less polar lipid membrane environment) while numerous fluorescence quenching assays can determine the depth of penetration of the peptide/fluorophore in the bilayer. Circular dichroism and Forster resonance energy transfer studies will elucidate peptide folding at the bilayer surface. Finally absorbance and fluorescence spectroscopy will be used in the bacterial membrane permeabilization assays, one of the proposed mechanisms of action of HDPs. The naturally occurring HDPs will first be examined with no sequence alterations and rationally designed analogues will be synthesized after this initial characterization. Sequence modifications focusing on the cationic amino acids in the sequence as well as modification of net charge/hydrophobicity of the HDPs will be the central experiments. HDP binding affinity and secondary structure data will be collected for a variety of lipid bilayer and detergent micelle compositions to determine lipid-based effects on HDP structure and mechanism of action. Other environmental factors that may impact mechanism of action including ionic strength and pH will also be investigated. Antimicrobial activity and bacterial membrane permeability assays will be performed using both gram positive and gram negative strains of bacteria. New, more broadly applicable assays for membrane permeabilization will also be developed. PUBLIC HEALTH RELEVANCE: This project involves determining the mechanism of action of a number of naturally occurring host defense peptides (HDPs), which are an essential component of the innate immune system in most higher organisms. The development of resistance to small-molecule and synthetic antimicrobials is a major public health concern, both in its annual death toll and its potential to develop "superbugs" or bacterial strains which are resistant to multiple last-line antibiotics. A full understanding of the structure-activity relationship in the mechanism of action of HDPs from varied host organisms is a key component necessary for the rational design of new antimicrobials with low potential for resistance development.

描述（由申请人提供）：本研究的总体目标是与膜相互作用抗菌肽的作用机制相关的化学和结构基序。许多细菌菌株中抗生素耐药性的增加每年都会产生巨大的影响，特别是在医院环境中，对于免疫系统受损的患者来说。随着越来越多的菌株对更多的抗生素产生耐药性，这就凸显了开发新的抗菌药物的必要性，这些抗菌药物的耐药性发展倾向较低。此类抗菌剂的一个潜在来源是天然存在的宿主防御肽（HDP），在整个进化链的多种生物体中都发现了这种肽，这表明细菌无法对这些分子产生足够的抵抗反应。 HDP 通常是短肽 (10-50 个氨基酸)，与宿主细胞相比具有高亲和力和高选择性，与细菌细胞表面结合。肽的作用机制仍然存在争议，但大多数 HDP 在与脂质双层结合后表现出一定程度的膜不稳定，这可能与作用机制有关。最近的研究还表明，暴露于 HDP 会诱导许多细菌转录反应，其中许多反应尚不完全清楚。从天然来源中分离出的 HDP 数量巨大，使得作用机制研究变得更加复杂，其中许多 HDP 可能根据宿主和宿主经常遇到的病原体而使用不同的机制。拟议研究的目标是重点研究针对不同宿主生物体和广谱抗菌活性而选择的几种天然衍生的 α-螺旋 HDP 的作用机制。对 α-螺旋 HDP 机制的更深入理解依赖于对这些肽的结构-活性关系的理解。此外，作为相对较小的肽，HDP 是实验上易于处理且有价值的生物物理模型，用于理解蛋白质-膜相互作用，并作为探索膜蛋白质折叠原理的模型系统。我们研究 HDP 行为的策略涉及使用天然和改变的肽序列合成肽，旨在检查特定化学部分在肽活性中的作用。这将通过合理地掺入天然存在的或合成的氨基酸来替换肽内一系列位点的相似残基、监测修饰肽与脂质双层的结合以及测试新设计的肽的抗菌活性来实现。将采用多种类型的光谱来表征肽中的膜结合和结构转变。荧光发射光谱是环境变化的敏感探针（即水性肽与极性较小的脂质膜环境结合），而许多荧光猝灭测定可以确定肽/荧光团在双层中的渗透深度。圆二色性和福斯特共振能量转移研究将阐明双层表面的肽折叠。最后，吸光度和荧光光谱将用于细菌膜透化测定，这是 HDP 的拟议作用机制之一。天然存在的 HDP 将首先在没有序列改变的情况下进行检查，并在初步表征后合成合理设计的类似物。集中于序列中的阳离子氨基酸的序列修饰以及HDP的净电荷/疏水性修饰将是中心实验。将收集各种脂质双层和去污剂胶束组合物的 HDP 结合亲和力和二级结构数据，以确定基于脂质对 HDP 结构和作用机制的影响。还将研究可能影响作用机制的其他环境因素，包括离子强度和 pH 值。将使用革兰氏阳性和革兰氏阴性细菌菌株进行抗菌活性和细菌膜渗透性测定。还将开发新的、更广泛适用的膜透化分析方法。公共健康相关性：该项目涉及确定许多天然存在的宿主防御肽（HDP）的作用机制，它们是大多数高等生物体先天免疫系统的重要组成部分。对小分子和合成抗菌药物产生耐药性是一个主要的公共卫生问题，无论是每年的死亡人数还是产生对多种最后一线抗生素具有耐药性的“超级细菌”或细菌菌株的潜力。充分了解来自不同宿主生物的 HDP 作用机制中的构效关系是合理设计低耐药性新型抗菌药物所必需的关键组成部分。

项目成果

期刊论文数量（10）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Painting proteins blue: ýý-(1-azulenyl)-L-alanine as a probe for studying protein-protein interactions.

将蛋白质涂成蓝色：××-(1-azulenyl)-L-丙氨酸作为研究蛋白质-蛋白质相互作用的探针。

DOI：
发表时间：
2013-01-18
期刊：
影响因子：
0
作者：
Moroz, Yurii S;Binder, Wolfgang;Nygren, Patrik;Caputo, Gregory A;Korendovych, Ivan V
通讯作者：
Korendovych, Ivan V

Impacts of Hydrophobic Mismatch on Antimicrobial Peptide Efficacy and Bilayer Permeabilization.

疏水错配对抗菌肽功效和双层通透性的影响。

DOI：
发表时间：
2023-11-14
期刊：
影响因子：
0
作者：
Meier, Steven;Ridgway, Zachary M;Picciano, Angela L;Caputo, Gregory A
通讯作者：
Caputo, Gregory A

Role of Cationic Side Chains in the Antimicrobial Activity of C18G.

阳离子侧链在 C18G 抗菌活性中的作用。

DOI：
发表时间：
2018-02-04
期刊：
影响因子：
0
作者：
Kohn, Eric M;Shirley, David J;Arotsky, Lubov;Picciano, Angela M;Ridgway, Zachary;Urban, Michael W;Carone, Benjamin R;Caputo, Gregory A
通讯作者：
Caputo, Gregory A

Investigation of the structure-activity relationship in ponericin L1 from Neoponera goeldii.

戈氏新珊瑚素 L1 构效关系的研究。

DOI：
发表时间：
2020-05
期刊：
影响因子：
0
作者：
Senetra, Alexandria S;Necelis, Matthew R;Caputo, Gregory A
通讯作者：
Caputo, Gregory A

Functional characterization of a melittin analog containing a non-natural tryptophan analog.

含有非天然色氨酸类似物的蜂毒肽类似物的功能表征。

DOI：
发表时间：
2015-07
期刊：
影响因子：
2.9
作者：
Ridgway, Zachary;Picciano, Angela L;Gosavi, Pallavi M;Moroz, Yurii S;Angevine, Christopher E;Chavis, Amy E;Reiner, Joseph E;Korendovych, Ivan V;Caputo, Gregory A
通讯作者：
Caputo, Gregory A